Transcription of the rabbit uteroglobin (UG) gene is regulated strongly by progesterone in the uterus, where estrogen has no (or little) effect, and weakly by glucocorticoids in the lung. As the appropriate steroid receptors are present in both tissues, additional factors may be involved in regulating uteroglobin transcription. Using a gel-shift assay, we have identified a trans-acting factor in nuclear extracts of progestational endometrium that specifically binds to the promoter region (UG203, -194/+9) of the UG gene. The major shift seen with progesterone-dominated endometrium was not present in non-target tissues (lung, liver), in HeLa cell nuclear extracts, or in estrogen-dominated endometrium. The amount of promoter-binding activity increased progressively with up to 5 days of treatment with progesterone, in parallel with the increase in UG mRNA concentration. Mixing experiments with non-specific nuclear extracts in which the specific shift was non seen, revealed an inhibitor of promoter binding. The inhibition was abolished when competitor extract proteins were heated (100 degrees celsius, 20 min) before addition to binding reactions. The inhibitor was also present in extracts of unstimulated endometrium and was reversed by the action of progesterone, with partial restoration of promoter-binding activity at 12 h after treatment. The results suggest that binding of a putative transcriptional factor to the UG promoter switches from negative to positive with the action of progesterone, and this switch shows some hormonal specificity as it did not occur with estrogen. The binding activity corresponds to the expression of the UG gene and this protein may be, therefore, a coordinately regulated transcription factor which regulates UG in both a tissue- and hormone- specific manner. The proposed studies are to purify the trans-activating factor using DNase I footprinting and DNA-affinity chromatography. The strategy is to then raise monospecific antibodies against the factor which can be used to screen an endometrial cDNA library to isolate the appropriate clone for the gene. The long term goals of the project are to study the interaction of two positive modulators of the gene (progesterone receptor, promoter-binding factor) and induction of gene expression. These interactions are central for understanding the regulation of progesterone mediated transcription in the mammalian uterus.
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| Rider, V; Peterson, C J (1991) Activation of uteroglobin gene expression by progesterone is modulated by uterine-specific promoter-binding proteins. Mol Endocrinol 5:911-20 |
